Molecular and Cellular Biochemistry

, Volume 377, Issue 1–2, pp 87–96 | Cite as

RETRACTED ARTICLE: TNF receptor-associated factor 6 regulates proliferation, apoptosis, and invasion of glioma cells

  • Zhang PengEmail author
  • Yue Shuangzhu
  • Jiang Yongjie
  • Zhang Xinjun
  • Liu Ying


Tumor necrosis factor receptor-associated factor 6 (TRAF6), which plays an important role in inflammation and immune response, is an essential adaptor protein for the NF-κB (nuclear factor κB) signaling pathway. Recent studies have shown that TRAF6 played an important role in tumorigenesis and invasion by suppressing NF-κB activation. However, up to now, the biologic role of TRAF6 in glioma has still remained unknown. To address the expression of TRAF6 in glioma cells, four glioma cell lines (U251, U-87MG, LN-18, and U373) and a non-cancerous human glial cell line SVG p12 were used to explore the protein expression of TRAF6 by Western blot. Our results indicated that TRAF6 expression was upregulated in human glioma cell lines, especially in metastatic cell lines. To investigate the role of TRAF6 in cell proliferation, apoptosis, invasion, and migration of glioma, we generated human glioma U-87MG cell lines in which TRAF6 was either overexpressed or depleted. Subsequently, the effects of TRAF6 on cell viability, cell cycle distribution, apoptosis, invasion, and migration in U-87MG cells were determined with 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) assay, flow cytometry analysis, transwell invasion assay, and wound-healing assay. The results showed that knockdown of TRAF6 could decrease cell viability, suppress cell proliferation, invasion and migration, and promote cell apoptosis, whereas overexpression of TRAF6 displayed the opposite effects. In addition, the effects of TRAF6 on the expression of phosphor-NF-κB (p-p65), cyclin D1, caspase 3, and MMP-9 were also probed. Knockdown of TRAF6 could lower the expression of p-p65, cyclin D1, and MMP-9, and raise the expression of caspase 3. All these results suggested that TRAF6 might be involved in the potentiation of growth, proliferation, invasion, and migration of U-87MG cell, as well as inhibition of apoptosis of U-87MG cell by abrogating activation of NF-κB.


Glioma TRAF6 Proliferation Apoptosis Invasion 


  1. 1.
    Farias-Eisner G, Bank AM, Hwang BY, Appelboom G, Piazza MA, Bruce SS, Sander Connolly E (2012) Glioblastoma biomarkers from bench to bedside: advances and challenges. Br J Neurosurg 26:189–194CrossRefPubMedGoogle Scholar
  2. 2.
    Walecki J, Tarasow E, Kubas B, Czemicki Z, Lewko J, Podgorski J, Sokol M, Grieb P (2003) Hydrogen-1 MR spectroscopy of the peritumoral zone in patients with cerebral glioma: assessment of the value of the method. Acad Radiol 10:145–153CrossRefPubMedGoogle Scholar
  3. 3.
    Ferguson SD (2011) Malignant gliomas: diagnosis and treatment. Dis Mon 57:558–569CrossRefPubMedGoogle Scholar
  4. 4.
    Rainov NG, Heidecke V (2011) Clinical development of experimental therapies for malignant glioma. Sultan Qaboos Univ Med J 11:5–28PubMedPubMedCentralGoogle Scholar
  5. 5.
    Yamanaka R, Kajiwara K (2012) Dendritic cell vaccines. Adv Exp Med Biol 746:187–200CrossRefPubMedGoogle Scholar
  6. 6.
    Jin HR, Jin SZ, Cai XF, Li D, Wu X, Nan JX, Lee JJ, Jin X (2012) Cryptopleurine targets NF-kappaB pathway, leading to inhibition of gene products associated with cell survival, proliferation, invasion, and angiogenesis. PLoS One 7:e40355CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Liu YQ, Hu XY, Lu T, Cheng YN, Young CY, Yuan HQ, Lou HX (2012) Retigeric acid B exhibits antitumor activity through suppression of nuclear factor-kappaB signaling in prostate cancer cells in vitro and in vivo. PLoS One 7:e38000CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Kang K, Lim JS (2012) Induction of functional changes of dendritic cells by silica nanoparticles. Immune Netw 12:104–112CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Prasad S, Yadav VR, Sung B, Reuter S, Kannappan R, Deorukhkar A, Diagaradjane P, Wei C, Baladandayuthapani V, Krishnan S et al (2012) Ursolic acid inhibits growth and metastasis of human colorectal cancer in an orthotopic nude mouse model by targeting multiple cell signaling pathways: chemosensitization with capecitabine. Clin Cancer Res 18:4942–4953CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Ge Y, Xu Y, Sun W, Man Z, Zhu L, Xia X, Zhao L, Zhao Y, Wang X (2012) The molecular mechanisms of the effect of dexamethasone and cyclosporin A on TLR4/NF-kappaB signaling pathway activation in oral lichen planus. Gene 508:157–164CrossRefPubMedGoogle Scholar
  11. 11.
    Jia L, Gopinathan G, Sukumar JT, Gribben JG (2012) Blocking autophagy prevents bortezomib-induced NF-kappaB activation by reducing I-kappaBalpha degradation in lymphoma cells. PLoS One 7:e32584CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Gupta SC, Sundaram C, Reuter S, Aggarwal BB (2010) Inhibiting NF-kappaB activation by small molecules as a therapeutic strategy. Biochim Biophys Acta 1799:775–787CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Tsuchiya Y, Asano T, Nakayama K, Kato T Jr, Karin M, Kamata H (2010) Nuclear IKKbeta is an adaptor protein for IkappaBalpha ubiquitination and degradation in UV-induced NF-kappaB activation. Mol Cell 39:570–582CrossRefPubMedGoogle Scholar
  14. 14.
    Tao T, Cheng C, Ji Y, Xu G, Zhang J, Zhang L, Shen A (2012) Numbl inhibits glioma cell migration and invasion by suppressing TRAF5-mediated NF-kappaB activation. Mol Biol Cell 23:2635–2644CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Tran NL, McDonough WS, Savitch BA, Fortin SP, Winkles JA, Symons M, Nakada M, Cunliffe HE, Hostetter G, Hoelzinger DB et al (2006) Increased fibroblast growth factor-inducible 14 expression levels promote glioma cell invasion via Rac1 and nuclear factor-kappaB and correlate with poor patient outcome. Cancer Res 66:9535–9542CrossRefPubMedGoogle Scholar
  16. 16.
    Sarkar D, Park ES, Emdad L, Lee SG, Su ZZ, Fisher PB (2008) Molecular basis of nuclear factor-kappaB activation by astrocyte elevated gene-1. Cancer Res 68:1478–1484CrossRefPubMedGoogle Scholar
  17. 17.
    Sung B, Pandey MK, Nakajima Y, Nishida H, Konishi T, Chaturvedi MM, Aggarwal BB (2008) Identification of a novel blocker of IkappaBalpha kinase activation that enhances apoptosis and inhibits proliferation and invasion by suppressing nuclear factor-kappaB. Mol Cancer Ther 7:191–201CrossRefPubMedGoogle Scholar
  18. 18.
    Ichikawa H, Takada Y, Murakami A, Aggarwal BB (2005) Identification of a novel blocker of I kappa B alpha kinase that enhances cellular apoptosis and inhibits cellular invasion through suppression of NF-kappa B-regulated gene products. J Immunol 174:7383–7392CrossRefPubMedGoogle Scholar
  19. 19.
    Schneider M, Zimmermann AG, Roberts RA, Zhang L, Swanson KV, Wen H, Davis BK, Allen IC, Holl EK, Ye Z et al (2012) The innate immune sensor NLRC3 attenuates Toll-like receptor signaling via modification of the signaling adaptor TRAF6 and transcription factor NF-kappaB. Nat Immunol 13:823–831CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Maruyama K, Kawagoe T, Kondo T, Akira S, Takeuchi O (2012) TRAF family member-associated NF-kappaB activator (TANK) is a negative regulator of osteoclastogenesis and bone formation. J Biol Chem 287:29114–29124CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Zhou F, Zhang X, van Dam H, Ten Dijke P, Huang H, Zhang L (2012) Ubiquitin-specific protease 4 mitigates toll-like/interleukin-1 receptor signaling and regulates innate immune activation. J Biol Chem 287:11002–11010CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Hamidi A, von Bulow V, Hamidi R, Winssinger N, Barluenga S, Heldin CH, Landstrom M (2012) Polyubiquitination of transforming growth factor beta (TGFbeta)-associated kinase 1 mediates nuclear factor-kappaB activation in response to different inflammatory stimuli. J Biol Chem 287:123–133CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Wang HJ, Ruan HJ, He XJ, Ma YY, Jiang XT, Xia YJ, Ye ZY, Tao HQ (2010) MicroRNA-101 is down-regulated in gastric cancer and involved in cell migration and invasion. Eur J Cancer 46:2295–2303CrossRefPubMedGoogle Scholar
  24. 24.
    Jacobs W, Mikkelsen T, Smith R, Nelson K, Rosenblum ML, Kohn EC (1997) Inhibitory effects of CAI in glioblastoma growth and invasion. J Neurooncol 32:93–101CrossRefPubMedGoogle Scholar
  25. 25.
    Meng QH, Zhou LX, Luo JL, Cao JP, Tong J, Fan SJ (2005) Effect of 7-hydroxystaurosporine on glioblastoma cell invasion and migration. Acta Pharmacol Sin 26:492–499CrossRefPubMedGoogle Scholar
  26. 26.
    Walsh MC, Kim GK, Maurizio PL, Molnar EE, Choi Y (2008) TRAF6 autoubiquitination-independent activation of the NFkappaB and MAPK pathways in response to IL-1 and RANKL. PLoS One 3:e4064CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Inubushi T, Kawazoe A, Miyauchi M, Kudo Y, Ao M, Ishikado A, Makino T, Takata T (2012) Molecular mechanisms of the inhibitory effects of bovine lactoferrin on lipopolysaccharide-mediated osteoclastogenesis. J Biol Chem 287:23527–23536CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Hartupee J, Li X, Hamilton T (2008) Interleukin 1alpha-induced NFkappaB activation and chemokine mRNA stabilization diverge at IRAK1. J Biol Chem 283:15689–15693CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Lim KH, Yang Y, Staudt LM (2012) Pathogenetic importance and therapeutic implications of NF-kappaB in lymphoid malignancies. Immunol Rev 246:359–378CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Baldwin AS (2012) Regulation of cell death and autophagy by IKK and NF-kappaB: critical mechanisms in immune function and cancer. Immunol Rev 246:327–345CrossRefPubMedGoogle Scholar
  31. 31.
    Ling J, Kumar R (2012) Crosstalk between NF-kB and glucocorticoid signaling: a potential target of breast cancer therapy. Cancer Lett 322:119–126CrossRefPubMedGoogle Scholar
  32. 32.
    Zhong L, Cao F, You Q (2012) Effect of TRAF6 on the biological behavior of human lung adenocarcinoma cell. Tumour Biol. doi: 10.1007/s13277-012-0543-8
  33. 33.
    Liu H, Zhang T, Ye J, Li H, Huang J, Li X, Wu B, Huang X, Hou J (2012) TNF receptor-associated factor 6 in advanced non-small cell lung cancer: clinical and prognostic implications. J Cancer Res Clin Oncol 138:1853–1863CrossRefPubMedGoogle Scholar
  34. 34.
    Chaudhry SI, Hooper S, Nye E, Williamson P, Harrington K, Sahai E (2012) Autocrine IL-1beta-TRAF6 signalling promotes squamous cell carcinoma invasion through paracrine TNFalpha signalling to carcinoma-associated fibroblasts. Oncogene. doi:  10.1038/onc.2012.91
  35. 35.
    Meng Q, Zheng M, Liu H, Song C, Zhang W, Yan J, Qin L, Liu X (2012) TRAF6 regulates proliferation, apoptosis, and invasion of osteosarcoma cell. Mol Cell Biochem 371:177–186CrossRefPubMedGoogle Scholar
  36. 36.
    Li YF, Xu XB, Chen XH, Wei G, He B, Wang JD (2012) The nuclear factor-kappaB pathway is involved in matrix metalloproteinase-9 expression in RU486-induced endometrium breakdown in mice. Hum Reprod 27:2096–2106CrossRefPubMedGoogle Scholar
  37. 37.
    Cock-Rada AM, Medjkane S, Janski N, Yousfi N, Perichon M, Chaussepied M, Chluba J, Langsley G, Weitzman JB (2012) SMYD3 promotes cancer invasion by epigenetic upregulation of the metalloproteinase MMP-9. Cancer Res 72:810–820CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Yang L, Zeng W, Li D, Zhou R (2009) Inhibition of cell proliferation, migration and invasion by DNAzyme targeting MMP-9 in A549 cells. Oncol Rep 22:121–126CrossRefPubMedGoogle Scholar
  39. 39.
    Jee BK, Park KM, Surendran S, Lee WK, Han CW, Kim YS, Lim Y (2006) KAI1/CD82 suppresses tumor invasion by MMP9 inactivation via TIMP1 up-regulation in the H1299 human lung carcinoma cell line. Biochem Biophys Res Commun 342:655–661CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Zhang Peng
    • 1
    Email author
  • Yue Shuangzhu
    • 1
  • Jiang Yongjie
    • 2
  • Zhang Xinjun
    • 3
  • Liu Ying
    • 2
  1. 1.Department of NeurosurgeryThe First Affiliated Hospital of Xinxiang Medical UniversityXinxiangChina
  2. 2.Department of Gynaecology and ObstetricsThe First Affiliated Hospital of Xinxiang Medical UniversityXinxiangChina
  3. 3.Department of UrologyThe First Affiliated Hospital of Xinxiang Medical UniversityXinxiangChina

Personalised recommendations